We have recently isolated several Ia+ cells lines, designated P388AD (adherent) and P388NA (nonadherent) from the P388 leukemia. Despite the similarity between the P388AD and P388NA cell lines with respect to Ia density, only the P388AD clones have been shown to induce syngeneic and allogeneic mixed leukocyte reactions, stimulate cytolytic T-cell activity against D and K H-2 antigens and induce cytolytic activity against TNP-coupled AD clones. Furthermore, antigen-pulsed P388AD but not P388NA clones induce proliferation of Ia-depleted primed T cells and long-term cultured T-cell lines. Last, preliminary information indicates that antigen-pulsed P388NA clones can induce T-cell proliferation in the presence of exogenous IL-1. On the basis of these data, we propose to use these cell lines to: (1)\determine the factors associated with the Ia+ P388AD cell lines which regulate the induction of antigen-independent T-cell proliferation in the syngeneic mixed leukocyte reaction (SMLR); (2)\analyze antigen presentation by P388AD and P388NA clones; (3)\determine if P388AD clones serve as accessory cells augmenting other in vitro immune responses; (4)\determine if proliferation signals in the SMLR represent a normal response during induction of antigen-specific T-cell proliferation; and (5)\characterize the physical interactions between antigen and Ia on the surface of P388AD clones. Of particular interest are experiments in which highly labeled MS2 coat protein will be fed to antigen-presenting cells, followed by treatment with bifunctional cross-linking reagents, specific precipitation with antisera to surface antigens (Ia, H-2D, Mac-1)\and tryptic peptide analysis of the antigen associated with the surface products. These experiments should provide new insights into the factors controlling T cell-accessory cell interactions and may also provide biochemical information on the nature of selective Ia-antigen fragment interactions on the antigen-presenting cell surface.